Chromosome-level de novo genome assembly of two conifer-parasitic wasps,Megastigmus duclouxiana and Megastigmus sabinae,reveals genomic imprints of adaptation to hosts

Conifers make up about one third of global forests but are threatened by seed parasitoid wasp species. Many of these wasps belong to the genus Megastigmus, yet little is known about their genomic background. In this study, we provide chromosome-level genome assemblies for two oligophagous conifer parasitoid species of Megastigmus, which represent the first two chromosome-level genomes of the genus. The assembled genomes of Megastigmus duclouxiana and M. sabinae are 878.48 Mb (scaffold N50 of 215.60 Mb) and 812.98 Mb (scaffold N50 of 139.16 Mb), respectively, which are larger than the genome size of most hymenopterans due to the expansion of transposable elements. Expanded gene families highlight the difference in sensory-related genes between the two species, reflecting the difference in their hosts. We further found that these two species have fewer family members but more single-gene duplications than polyphagous congeners in the gene families of ATP-binding cassette transporter (ABC), cytochrome P450 (P450) and olfactory receptors (OR). These findings shed light on the pattern of adaptation to a narrow spectrum of hosts in oligophagous parasitoids. Our findings suggest potential drivers underlying genome evolution and parasitism adaptation, and provide valuable resources for understanding the ecology, genetics and evolution of Megastigmus, as well as for the research and biological control of global conifer forest pests.     

Divergence in gene expression related to variation in host specificity of an ectomycorrhizal fungus

Ectomycorrhizae are formed by mutualistic interactions between fungi and the roots of woody plants. During symbiosis the two organisms exchange carbon and nutrients in a specific tissue that is formed at the contact between a compatible fungus and plant. There is considerable variation in the degree of host specificity among species and strains of ectomycorrhizal fungi. In this study, we have for the first time shown that this variation is associated with quantitative differences in gene expression, and with divergence in nucleotide sequences of symbiosis-regulated genes. Gene expression and sequence evolution were compared in different strains of the ectomycorrhizal fungus Paxillus involutus; the strains included Nau, which is not compatible with birch and poplar, and the two compatible strains Maj and ATCC200175. On a genomic level, Nau and Maj were very similar. The sequence identity was 98.9% in the 16 loci analysed, and only three out of 1075 genes analysed by microarray-based hybridizations had signals indicating differences in gene copy numbers. In contrast, 66 out of the 1075 genes were differentially expressed in Maj compared to Nau after contact with birch roots. Thirty-seven of these symbiosis-regulated genes were also differentially expressed in the ATCC strain. Comparative analysis of DNA sequences of the symbiosis-regulated genes in different strains showed that two of them have evolved at an enhanced rate in Nau. The sequence divergence can be explained by a decreased selection pressure, which in turn is determined by lower functional constraints on these proteins in Nau as compared to the compatible strains.     

Genetic innovations: Transposable element recruitment and de novo formation lead to the birth of orphan genes in the rice genome

Orphan genes are genetic innovations that lack homologs in other lineages. Orphan genes can rapidly originate and become substantially functional, yet the mechanisms underlying their origins are still largely unknown in plants. Here, we investigated the origin of orphan genes in the Oryza sativa ssp. japonica “Nipponbare” genome using genome‐wide comparisons with 10 closely related Oryza species. We identified a total of 37 orphan genes in the Nipponbare genome that show short sequence lengths, elevated GC content, and absence of introns. Interestingly, half of the identified orphan genes originated by way of a distinctive mechanism that involved the generation of new coding sequences through independent and rapid divergence within the inserted transposable element. Our results provide valuable insight into genetic innovations in the model rice genome that formed on a very short timescale.     

Selection of reference genes for qRT‐PCR and expression analysis of high‐altitude‐related genes in grassland caterpillars (Lepidoptera: Erebidae: Gynaephora) along an altitude gradient

Changes in gene expression patterns can reflect the adaptation of organisms to divergent environments. Quantitative real‐time PCR (qRT‐PCR) is an important tool for ecological adaptation studies at the gene expression level. The quality of the results of qRT‐PCR analysis largely depends on the availability of reliable reference genes (RGs). To date, reliable RGs have not been determined for adaptive evolution studies in insects using a standard approach. Here, we evaluated the reliability of 17 candidate RGs for five Gynaephora populations inhabiting various altitudes of the Tibetan Plateau (TP) using four independent (geNorm, NormFinder, BestKeeper, and the deltaCt method) and one comprehensive (RefFinder) algorithms. Our results showed that EF1‐α, RPS15, and RPS13 were the top three most suitable RGs, and a combination of these three RGs was the most optimal for normalization. Conversely, RPS2, ACT, and RPL27 were the most unstable RGs. The expression profiles of two target genes (HSP70 and HSP90) were used to confirm the reliability of the chosen RGs. Additionally, the expression patterns of four other genes (GPI, HIF1A, HSP20, and USP) associated with adaptation to extreme environments were assessed to explore the adaptive mechanisms of TP Gynaephora species to divergent environments. Each of these six target genes showed discrepant expression patterns among the five populations, suggesting that the observed expression differences may be associated with the local adaptation of Gynaephora to divergent altitudinal environments. This study is a useful resource for studying the adaptive evolution of TP Gynaephora to divergent environments using qRT‐PCR, and it also acts as a guide for selecting suitable RGs for ecological and evolutionary studies in insects.     

褐飞虱热胁迫下内参基因的筛选及热激蛋白基因表达分析（英文）

【目的】褐飞虱Nilaparvata lugens(Stl)是为害水稻的重要害虫之一,温度是影响其暴发、迁飞的主要环境因子之一。本研究旨在探讨研究褐飞虱对高温胁迫适应性的热激蛋白基因表达调控模式。【方法】分别以不同的高温(30℃~40℃)处理褐飞虱雌、雄虫1 h和2 h,利用荧光定量PCR技术检测其体内的β-actin 1,β-actin2,β-actin3,28S rRNA,18S rRNA和α-2-tubulin 6个内参基因的表达量,用geN orm和BestK eeper软件分析确定最稳定表达的内参基因,并检测热胁迫后hsp70和hsp90基因在处理褐飞虱成虫体内的表达模式。【结果】geN orm软件分析结果表明,热胁迫后褐飞虱内参基因稳定性在雌虫体内为:β-actin1=β-actin328S rRNAα-2-tubulin18S rRNAβ-actin2;在雄虫体内为:β-actin1=β-actin3α-2-tubulin28S rRNA18S rRNAβ-actin2。BestK eeper软件分析结果显示,在热胁迫的雌、雄虫体内β-actin1均最稳定,18S rRNA次之,β-actin2最不稳定。两种软件分析结果基本一致。以β-actin1为校正内参基因,荧光定量PCR分析hsp70和hsp90在不同热胁迫条件下的表达模式,结果表明,各高温处理下hsp70表达量与对照26℃下的表达量没有显著性差异;而hsp90基因表达模式表现为被高温诱导上调表达,在雌、雄虫体内表达量达到最高的处理条件分别为40℃和38℃处理2 h。【结论】β-actin1基因可以作为热胁迫下褐飞虱雌雄虫体内基因表达模式分析的校正内参基因使用。褐飞虱hsp90基因能被高温诱导表达,该基因可能在褐飞虱适应热胁迫过程中起着重要的作用。     

mRNA levels of four tomato (Lycopersicon esculentum Mill. L.) genes related to fluctuating plant and soil water status

Expression of the genes le20, ni3212, le25 and lcyp2 was quantified in relation to soil and plant water status for pot-grown tomato (Lycopersicon esculentum Mill. cv. Ailsa Craig) in a replicated trial in a partially environmentally controlled glasshouse. Leaf water potential, relative water content, stomatal conductance and gravimetric soil water content were measured at 0800,1200 and 1600 h on three days during the imposition of three different watering regimes. Paired leaf samples were analysed for mRNA. Plant-to-plant variability was quantified and partitioned. le20 mRNA was found in all leaves and responded both to short-term diurnal fluctuations in water status and to more severe stress, though with reduced sensitivity to the greatest stresses. An overnight decline in le20 mRNA was independent of soil or plant water status. The pattern of expression for ni3212 was similar though ni3212 mRNA was much more abundant and the overnight drop in expression was not significant. le25 mRNA was only detected where leaf water potential was below ?0.9 MPa and increased linearly with further decline in leaf water potential. lcyp2 was constitutively expressed with a diurnal pattern characterized by a sharp depression at 1600 h but with expression being enhanced by increasing water deficit at all times. The control and significance of the four genes are discussed in relation to the proposed function of their respective gene products.     

Identification of genes related to Parkinson's disease using expressed sequence tags.

In a search for novel target genes related to Parkinson's disease (PD), two full-length cDNA libraries were constructed from a human normal substantia nigra (SN) and a PD patient's SN. An analysis of the gene expression profiles between them was done using the expressed sequence tags (ESTs) frequency. Data for the differently expressed genes were verified by quantitative real-time RT-PCR, immunohistochemical analysis and a cell death assay. Among the 76 genes identified with a significant difference (P > 0.9), 21 upregulated genes and 13 downregulated genes were confirmed to be differentially expressed in human PD tissues and/or in an MPTP-treated mice model by quantitative real-time RT-PCR. Among those genes, an immunohistochemical analysis using an MPTP mice model for alpha-tubulin including TUBA3 and TUBA6 showed that the protein levels are downregulated, as well as the RNA levels. In addition, MBP, PBP and GNAS were confirmed to accelerate cell death activity, whereas SPP1 and TUBA3 to retard this process. Using an analysis of ESTs frequency, it was possible to identify a large number of genes related to human PD. These new genes, MBP, PBP, GNAS, SPP1 and TUBA3 in particular, represent potential biomarkers for PD and could serve as useful targets for elucidating the molecular mechanisms associated with PD.     

The draft genome of a wild barley genotype reveals its enrichment in genes related to biotic and abiotic stresses compared to cultivated barley

Wild barley (Hordeum spontaneum) is the progenitor of cultivated barley (Hordeum vulgare) and provides a rich source of genetic variations for barley improvement. Currently, the genome sequences of wild barley and its differences with cultivated barley remain unclear. In this study, we report a high‐quality draft assembly of wild barley accession (AWCS276; henceforth named as WB1), which consists of 4.28 Gb genome and 36 395 high‐confidence protein‐coding genes. BUSCO analysis revealed that the assembly included full lengths of 95.3% of the 956 single‐copy plant genes, illustrating that the gene‐containing regions have been well assembled. By comparing with the genome of the cultivated genotype Morex, it is inferred that the WB1 genome contains more genes involved in resistance and tolerance to biotic and abiotic stresses. The presence of the numerous WB1‐specific genes indicates that, in addition to enhance allele diversity for genes already existing in the cultigen, exploiting the wild barley taxon in breeding should also allow the incorporation of novel genes. Furthermore, high levels of genetic variation in the pericentromeric regions were detected in chromosomes 3H and 5H between the wild and cultivated genotypes, which may be the results of domestication. This H. spontaneum draft genome assembly will help to accelerate wild barley research and be an invaluable resource for barley improvement and comparative genomics research.     

Modulation of enzyme activities and expression of genes related to primary and secondary metabolism in response to UV-B stress in cucumber (Cucumis sativus L.)

Abstract

Exposure to UV-B at ambient or enhanced levels is known to trigger a variety of responses in all living organisms, including higher plants. Here we show that in Cucumis sativus L. UV-B radiation affects enzyme activity of key oxydative pentose phosphate pathway (OPPP) enzymes glucose-6-phosphate dehydrogenase (G6P-DH) and 6-phosphogluconate dehydrogenase (6-PGlu-DH), of key phenolic compounds enzyme phenylalanine ammonia lyase (PAL) as well as erythrose-4-phosphate, tryptophan and tyrosine levels. Furthermore, we found an increased activity of antioxidant enzymes such as peroxidase (POX) and catalase (CAT) in treated plants, with respect to the controls. In order to confirm the biochemical results, we isolated total RNA from both controls and UV-B treated plants to be used for gene expression analysis. We demonstrated that UV-B increases the gene expression level of peroxidase (POX), catalase (CAT) and phenylalanine ammonia lyase (PAL). Finally, our results are useful for understanding protective strategies against UV-B radiation and for elucidating what components are involved in stress-induced signals within the plant.     

Effects of branched-chain volatile fatty acids on lactation performance and mRNA expression of genes related to fatty acid synthesis in mammary gland of dairy cows

Branched-chain volatile fatty acids (BCVFA) supplements could promote lactation performance and milk quality by improving ruminal fermentation and milk fatty acid synthesis. This study was conducted to evaluate the effects of BCVFA supplementation on milk performance, ruminal fermentation, nutrient digestibility and mRNA expression of genes related to fatty acid synthesis in mammary gland of dairy cows. A total of 36 multiparous Chinese Holstein cows averaging 606±4.7 kg of BW, 65±5.2 day in milk (DIM) with daily milk production of 30.6±0.72 kg were assigned to one of four groups blocked by lactation number, milk yield and DIM. The treatments were control, low-BCVFA (LBCVFA), medium-BCVFA (MBCVFA) and high-BCVFA (HBCVFA) with 0, 30, 60 and 90 g BCVFA per cow per day, respectively. Experimental periods were 105 days with 15 days of adaptation and 90 days of data collection. Dry matter (DM) intake tended to increase, but BW changes were similar among treatments. Yields of actual milk, 4% fat corrected milk, milk fat and true protein linearly increased, but feed conversion ratio (FCR) linearly decreased with increasing BCVFA supplementation. Milk fat content linearly increased, but true protein content tended to increase. Contents of C4:0, C6:0, C8:0, C10:0, C12:0, C14:0 and C15:0 fatty acids in milk fat linearly increased, whereas other fatty acids were not affected with increasing BCVFA supplementation. Ruminal pH, ammonia N concentration and propionate molar proportion linearly decreased, but total VFA production and molar proportions of acetate and butyrate linearly increased with increasing BCVFA supplementation. Consequently, acetate to propionate ratios linearly increased. Digestibilities of DM, organic matter, CP, NDF and ADF also linearly increased. In addition, mRNA expressions of peroxisome proliferator-activated receptor γ, sterol regulatory element-binding factor 1 and fatty acid-binding protein 3 linearly increased, mRNA expressions of acetyl-coenzyme A carboxylase-α, fatty acid synthase and stearoyl-CoA desaturase quadratically increased. However, lipoprotein lipase mRNA expression was not affected by treatments. The results indicated that lactation performance and milk fat synthesis increased with BCVFA supplementation by improving ruminal fermentation, nutrient digestibility and mRNA expressions of genes related to milk fat synthesis.